This invention relates to novel ultrasonic contrast agents, their production and use, more particularly such agents which are useful as left heart contrast agents.
The ultrasonic examination of organs (sonography) is a diagnostic method that has been popular and has been practiced for a number of years. Ultrasonic waves in the megahertz range (above 2 megahertz with wavelengths of between 1 and 0.2 mm) are reflected on interfaces of various types of tissue. The thus-produced echoes are amplified and made visible. Of special significance in this connection is the examination of the heart by this method, called echocardiography. See Haft, J. I. et al.: "Clinical Echocardiography", Futura, Mount Kisco, New York 1978; Kohler, E., "Klinische Echokardiographie" [Clinical Echocardiography], Enke, Stuttgart 1979; Stefan, G., et al.: "Echokardiographie" [Echocardiography], Thieme, Stuttgart-New York 1981; G. Biamino, L. Lange: "Echokardiographie", Hoechst AG, 1983.
Since liquids, including blood, yield ultrasonic contrast only if there are differences in density with respect to the surrounding area, possibilities have been explored for making the blood and its flow visible for ultrasonic analysis, which can be achieved by the addition of extremely small gas bubbles (microbubbles).
Several methods are known in the literature for the preparation and stabilization of microbubbles. They can be produced, for example, prior to injection into the blood stream by vigorous shaking or agitation of solutions, such as saline solutions , colorant solutions, or previously drawn blood. Although ultrasonic contrast imaging has been achieved by these methods, they have serious disadvantages which manifest themselves in poor reproducibility, widely fluctuating size of the microbubbles and, due to the concurrent production of a proportion of visible, larger bubbles, the risk of embolism.
These disadvantages have been overcome, in part, by manufacturing methods such as, for example, by U.S. Pat. No. 3,640,271 wherein small bubbles of reproducible size are generated by filtration or by the use of an electrode device using direct current. Although this method has the advantage of being able to produce small gas bubbles of reproducible size, it has the disadvantage of considerable technical costs.
U.S. Pat. No. 4,276,885 describes the production of microbubbles of a definite size, which are protected from coalescence by being surrounded by a gelatin envelope. However, the finished microbubbles disclosed therein must be stored in a "frozen" condition, for example, at refrigerator temperature and prior to use they must be brought to body temperature.
U.S. Pat. No. 4,264,251 discloses the manufacture and use of gas-filled microbubbles with a solid surrounding wall of a saccharide, which bubbles can be filled with a pressurized gas. If the microbubbles are at ambient pressure, they can be utilized as ultrasonic contrast media. If they have increased internal pressure, the microbubbles can be used for blood pressure determination. Although the storage of these solid microbubbles does not present a problem, the technical expenditure in their manufacture is a considerable cost factor.
Commonly assigned U.S. Pat. No. 4,466,442 claims a method of increasing the useful lifetime and the amount of microbubbles of a size less than 50 .mu.m produced by mechanical agitation of an aqueous liquid employed as an ultrasonic contrast medium employing a liquid containing dissolved therein a tenside which reduces the surface tension of the liquid and a compound which raises the viscosity of the liquid.
U.S. Pat. No. 4,572,203 claims a method of ultrasonic imaging for use in medical procedures which comprises injecting biodegradable, metal-containing microparticles, or microparticles comprising an amino acid matrix containing air, glass, graphite, nitrogen, carbon dioxide, metal flakes, magnetite, magnetic iron oxides or carbonyl iron, or sonicated microbubbles into an animal or human to thereby alter the acoustic properties of an area to be imaged, and then ultrasonically scanning the area so as to obtain an image.
Commonly assigned U.S. Pat. No. 4,681,119 claims a method wherein microbubbles are formed in a body liquid or carrier liquid, for example within the genitourinary or digestive systems in order to alter the transmission characteristics thereof to electromagnetic and sonic waves transmitted therethrough, by dissolving therein a solid particulate material, preferably as a suspension in a carrier liquid in which the particulate material is at least temporarily stable, the particles of which are substantially free of microbubbles and have a plurality of gas-filled voids communicating with the surface of the particles and providing nuclei for microbubble formation and the ratio of the mass of the particles to the volume of gas in the voids is sufficient to render the liquid in which the particulate material is dissolved supersaturated with respect to the gas in the voids in the area of the liquid surrounding the microbubbles when they are formed.
The following references were cited (along with DE-C-3-637 926) in the European Search Report issued in the European application corresponding to application Ser. No. 07/333,408.
EP 122 624 and EP 123 235 are European patents corresponding to the ancestor applications of application Ser. No. 07/333,408, which were combined as U.S. application Ser. No. 06/600,691, filed Apr. 16, 1984, which was abandoned in favor of continuation-in-part application Ser. No. 06/917,574, filed Oct. 10, 1986, which was abandoned in favor of application Ser. No. 07/370,140, filed Jun. 10, 1989, which will be abandoned in favor of the present application.
In parent U.S. application Ser. No. 07/333,408 (and EP 122,624), an image enhancing ultrasonic contrast agent containing microparticles and small gas bubbles is described which is suitable for enhancing, after intravenous administration and passage through the lungs, contrast imaging of the left side of the heart, of the myocardium, as well as other organs, such as the liver, the spleen, and the kidneys. The application cites fatty acids ["saturated or unsaturated (C.sub.4 -C.sub.20)-fatty acids"] as being suitable for use in the production of the microparticles. Confirmation is provided by surfactant esters and salts thereof, such as, for example, ascorbyl palmitate or sucrose monopalmitate. Compared to the preferred surfactants disclosed herein, these specific surfactants are relatively quickly decomposed in the formulation even when stored under normal conditions (25.degree. C.), which is disadvantageous for a commercial preparation and its purity requirements.
The related imaging agent of U.S. Pat. No. 4,442,843 similarly decomposes when stored.
DE-C-3-637 926, which corresponds to U.S. Pat. No. 4,684,479, cited above, describes the use of known agents, e.g., agents of EP 122 624, in an ultrasonic manometry process, which is unrelated to imaging.
The risks inherent in the use of an ultrasonic contrast medium are based in part on the size and number of any solid particles present therein as well as the size of the microbubbles therein.
Commonly assigned U.S. Pat. No. 4,442,843 (which corresponds to European Patent Application 81730118.7, Publication No. 52575), describes the production of microbubbles said to exhibit these required properties. For the manufacture of microbubbles according thereto, aggregates of particles of a solid, crystalline compound, e.g., galactose, which themselves lack microbubbles but which contain a gas which is adsorbed on the surface of the particles, occluded in cavities between the particles or is present in intracrystalline cavities, are suspended in a liquid vehicle just prior to use, thereby forming a suspension of undissolved particles and a small amount of microbubbles produced by particles dissolving in the liquid vehicle. This suspension is then injected into a blood stream within 10 minutes, where the major proportion (about 80%) of the microbubbles are formed when the undissolved aggregates in the suspension dissolve in the blood.
Although it is stated in these documents that the resultant suspension is suitable, after injection into a peripheral vein, not only for ultrasonic visualization of the right side of the heart but also, after passing through the lungs, for visualization of the left side of the heart and the blood and its flow at that location, it has not been possible to achieve reproducible and practically acceptable levels of left heart visualization employing the water soluble microbubble precursors disclosed therein. For example, when a contrast medium consisting of 400 mg. of aggregates of galactose microparticles (&lt;8 .mu.m; 2.5 .mu.m mean size) in 1 ml of a 20% aqueous lactose solution was injected into a peripheral vein, it did not evoke practically significant ultrasonic echoes in the left portion of the heart, apparently because the lifetime of the microbubbles formed in the blood was too short to permit them to pass through the capillaries of the lungs. Conversely, from two specific production lots of product obtained from the same manufacturer, batches of water soluble microbubble precursor according to that patent were produced, one formed from galactose and one from maltose, which, when suspended in 1 g/ml of a 90% aqueous glucose (5% solution)/10% propylene glycol mixture and injected into a peripheral vein, evoked significant left heart contrast. However, no other lots of galactose or maltose from the same or other manufacturer did so. No explanation for this was ever found.
To ultrasonically visualize the left heart, a blood vessel flowing therefrom or an organ receiving blood therefrom, either the microbubbles or an ultrasonic contrast agent capable of producing such microbubbles in situ in the blood must be injected into a blood vessel leading from the lungs to the heart, or into an artery (a highly undesirable procedure) or the microbubbles or a microbubble precursor which produces microbubbles in situ in the blood after injection into a peripheral vein must survive the passage through the lungs. The latter alternative requires that the size of the microbubbles or the microbubble precursor be less than 12, preferably less than 8 and most preferably in the range of about 1-3 micrometers. Such small microbubbles and microbubble precursors when produced conventionally by prior art methods have an extremely short lifespan. Therefore, the chances of their surviving a passage through the lungs is highly unlikely.
EP-A-77752 discloses the preparation of a liquid mixture for use as a contrast medium consisting, in turn, of a mixture of a tenside or an aqueous solution of the tenside, and an aqueous, viscous carrier liquid.
Right heart ultrasonic contrasting for a period of time long enough for imaging thereof can readily be achieved with the water soluble microbubble precursors of U.S. Pat. No. 4,442,843 by injecting a suspension thereof in a suitable liquid carrier into a peripheral vein as a bolus, which travels essentially unmixed with blood until it reaches the right heart, where the suspension is rapidly mixed with the blood therein by hydrodynamic effects. Virtually complete loss of ultrasonic image contrast ordinarily occurs beyond the pulmonary artery. Consequently, the use of an unmodified water soluble microbubble precursor of the type disclosed in U.S. Pat. No. 4,442,843 is not feasible for left heart contrast.
U.S. Pat. No. 4,442,843 states several factors that are said to improve left heart contrast using the water soluble microbubble precursors disclosed therein. These factors may increase the very small number of microbubbles which occasionally cross the lungs during use of such materials as right heart contrast agents, but in no case does such use reproducibly result in a number of microbubbles which is of practical use for left heart contrast purposes. These factors have been found to be irrelevant for use of the improved microbubble precursors of this invention by which highly practical and reproducible left heart contrast can be achieved.
For example, U.S. Pat. No. 4,442,843 states that although for general purposes aggregates in the 30-50 micrometer size range are preferred, for traversing the lungs to image the left heart aggregates of 125 micrometer average size is preferable. However, it has since been found that these aggregates rapidly disassociate into discrete microparticles either in the presence of the liquid carrier employed therewith or in the blood, apparently as a result of the van der Waals forces, which initially caused the microparticles to coalesce into aggregates, being overcome. Therefore, increasing aggregate size does not achieve practical left heart contrasting with solid microbubble precursors of that application.
That patent also states that for left heart imaging, optimum solubility of the solid precursor is in the range of 1 mole/liter and therefore galactose is superior to the other saccharides. We have since found that notwithstanding its lower solubility compared to other saccharides, particles of galactose in the 1-3 .mu.m range dissolve almost instantaneously in blood and therefore are no better than other saccharides for practical left heart contrasting, unless modified in accordance with this invention.
That patent also states that bubble production can be improved, inter alia. by the inclusion of a small amount of surfactant in the solid precursor or its carrier liquid. However, it has been found that conventional surfactants which have an HLB value above about 20, e.g., sodium lauryl sulfate, sodium dodecylbenzenesulfonate and the commercially available nonionic surfactants, e.g., the Pluronics, do not enhance practical left heart contrast. In fact, because such surfactants should facilitate solution of microparticles and microbubbles in blood, their use in conjunction with ultrasonic contrast agents intended for left heart imaging would appear to be contraindicated. Therefore, even with larger aggregate and smaller particle size and the use of a surfactant in the particles or in the liquid carrier, the water soluble microbubble precursors of U.S. Pat. No. 4,442,843 are not suitable for left heart ultrasonic imaging.
For the foregoing reasons, satisfactory left heart visualization with the prior art ultrasonic contrast agents, including the microbubble precursors disclosed in U.S. Pat. No. 4,442,843 could not be achieved in a predictable and reproducible manner.
The state of the prior art thus far permits the manufacture of ultrasonic contrast agents which exhibit only some of the following required or desired properties:
(1) elimination of the risk of embolism PA0 (2) reproducibility; PA0 (3) adequately long stability; PA0 (4) ability to pass through the lungs, for example, to obtain ultrasonic contrasting of the left portion of the heart and organs receiving blood therefrom; PA0 (5) ability to pass through capillaries; PA0 (6) sterility and freedom from pyrogens; PA0 (7) easily produced with feasible financial expenditures; and PA0 (8) storability without problems or special conditions.
small gas bubbles (size and number), PA1 solid particles (size and number);
It is an object of the present invention to provide a contrast agent suitable for altering the transmission characteristics of an aqueous liquid to an electromagnetic or elastic wave transmitted therethrough.
It is a further object of the present invention to provide a contrast agent for ultrasonic diagnostics which is capable, after intravenous administration, of ultrasonic imaging of the blood and its flow conditions not only in the right side of the heart but also, after passing through the capillary bed of the lung, in the left side of the heart and, consequently, in other organs, such as the myocardium, the liver, the spleen and the kidneys.
It is another object to provide ultrasonic contrasting methods employing these contrast agents. Other objects will be apparent to those skilled in the art to which this invention pertains.